Proteolysis is increasingly believed to regulate specific aspects of cellular function including interactions between cells. The regulatory influence of proteolysis on neural processes is the focus of this proposal. We have developed methods which now permit us to reliably study the mechanisms of axonal protein degradation within a specific neuron, the retinal ganglion cell, throughout development and during neuropathological states. Briefly, segments of mouse optic nerve containing axonally transported proteins isotopically labeled in vivo are incubated with protein synthesis inhibitors in vitro and the rate of amino acid release from protein, reflecting protein degradation, is measured. Complementary studies of proteolysis in glial cells can be made by slightly modifying this technique. Conducting the analysis at the level of organotypic organization preserves much of the physiological integrity of the proteolytic process and also allows the role of proteolysis in cell-cell interactions to be explored. Using these methods we have obtained evidence which points to important regulatory functions of axonal proteolysis during axonal degeneration and myelinogenesis. Our major objective in this proposal is to define these functions in detail. Specifically, we plan to further characterize the molecular components of the proteolytic machinery within axons and glial cells of the normal adult optic nerve using specific chemical probes in vitro. Comparably detailed analyses of proteolysis will be made during the evolution of Wallerian degeneration and of axonal degeneration in experimental polyneuropathies. Similarly, we will characterize the changes in axonal and glial proteolysis during normal myelinogenesis and assess their functional relationship to the myelination process by also analyzing mice with different gentic or chemically-induced defects of myelin development. Finally, we plan to develop means to experimentally manipulate specific proteolytic events for the purpose of modifying these neural phenomena in vivo. The information sought in these experiments is directly pertinent to the molecular mechanisms underlying many human neuropathies and certain neurodegenerative states as well as developmental brain dysfunction leading to mental retardation. The results may point to specific therapeutic approaches to these problems.